The prevention of undesired cell death has become an important goal for pharmacologic intervention in a variety of clinical settings. Profound disability can result from apoptotic cell death and tissue injury due to ischemia, chemotherapeutic agents, ionizing radiation, or hyperthermia. Recently, a small molecule, 2-(2-imino-4,5,6,7-tetrahydro-benzothiazol-3-yl)-1-p-tolyl-ethanone hydrobromide (PFT-α; 1),
was originally identified from a broad screen of 10,000 compounds to inhibit cell death from gamma radiation, Komarov, P. G.; et al., Science 1999, 285, 1733-1737. In addition, Komarov's results indicated that this compound was shown to protect mice from lethal genotoxic stress. In addition, it was discovered that PFT-α was also shown to protect focal cortical ischemic injury and neuronal excitotoxic damage, Culmsee, C., et al., J. Neurochem. 2001, 77, 220-228, and Culmsee, C. et al., Brain Res Mol Brain Res. 2001, 87, 257-262. The protection provided by PFT-α was attributed to inhibition of p53 transactivation. Supporting evidence has included diminished nuclear accumulation with decreased p53 DNA binding activity, decreased caspase activity and suppression of mitochondrial dysfunction. Furthermore, the transcription of apoptosis-associated gene products p53, Bax and p21 was inhibited, Proietti De Santis, L., et al., DNA Repair 2003, 2, 891-900.
However, the activity of PFT-α is not limited to inhibiting the transactivation of p53. This small molecule has been found to also suppress the heat shock and glucocorticoid signaling pathways Komarova, E. A., et al., J Biol Chem. 2003, 278, 15465-15468. Glucocorticoid-induced cell death is independent of p53 Clarke, A. R., et al., Nature 1993, 362, 849-852. This pathway, however, also induces the transcription and activity of the proapoptotic BH3 (Bcl-2 Homology)-only protein p53-upregulated modulator of apoptosis (PUMA) Han, J.-W.; Flemington, C. et al., Proc Natl Acad Sci USA. 2001, 98, 11318-11323. When this death-inducing signal reaches the mitochondria, a series of cell-death related events ensues. The inner mitochondrial membrane loses its potential and cytochrome c is released into the cytoplasm, where it can associate with ATP, apoptosis-activating factor-1, and procaspase-9. This apoptosome complex cleaves procaspase-9 to caspase-9, which in turn cleaves procaspase-3, initiating the cascade of protease activation in the execution phase of apoptosis Li, P., et al., Cell 1997, 91, 479-489. and Zou, H., et al., Cell 1997, 90, 405-413.
PFT-α has been reported to diminish p53-dependent and independent mitochondria mediated cell death in vitro and in vivo, Komarov, P. G.; et al., Science 1999, 285, 1733-1737 and Komarova, E. A., et al., J Biol Chem. 2003, 278, 15465-15468. The multiplicity of molecular pathways that are reportedly influenced by PFT-α suggests that the cytoprotective effect could be further optimized. Furthermore, PFT-α is not stable under physiologic conditions and spontaneously undergoes ring closure to form the imidazo[2,1-b]benzothiazole (IBT) 2, Singh, A.; Mohan, J., et al., Indian J. Chem., Sect. B 1976, 14B, 997-998.
In spite of recent reports by Zhu, X., et al., J Med Chem. 2002, 45, 5090-5097, and Pietrancosta, N., et al., Bioorg Med Chem Lett. 2005, 15, 1561-1564, describing a few of the compounds prepared here, the biologically active form of PFT-α (open versus closed ring) has not previously been formally determined. Elucidation of the active ring structure would allow investigation of further chemical modifications that may enhance the potency of the compound. In addition, alternative ring structures, such as the quaternary salts recently reported from our laboratories, may also provide greater potency Barchechath, S. D., et al., Bioorg Med Chem Lett. 2005, 15, 1785-1788.
Thus, we report the synthesis and structure activity relationship of novel derivatives and analogs of PFT-α, and corresponding closed ring counterparts, with enhanced potency and stability. To determine activity, we used a p53-independent assay of cell death wherein mouse thymocytes were treated with dexamethasone in the presence or absence of test compounds. In addition, we confirmed the activity of the most potent compounds in a p53-dependent apoptosis assay.
In addition, there is a need for novel, potent, and selective agents to prevent detrimental effects upon cells due to DNA damage, such as caused by chemotherapy, radiation, ischemic event, including ischemia-reperfusion injury and organ transplantation, and the like. There is also a need for pharmacological tools for the further study of the physiological processes associated with intracellular DNA damage.
A continuing need exists for compounds that can protect mammalian cells from the damaging effects of chemotherapy and irradiation, or in other situations in which it is desirable to protect tissue from the consequences of clinical or environmental stress.